Changes in S-(2-succinyl)cysteine and advanced glycation end-products levels in mouse tissues associated with aging

Katsuta, Nana; Takahashi, Himeno; Nagai, Mime; Sugawa, Hikari; Nagai, Ryoji

doi:10.1007/s00726-022-03130-y

Original Article
Published: 15 February 2022

Changes in S-(2-succinyl)cysteine and advanced glycation end-products levels in mouse tissues associated with aging

Nana Katsuta¹,
Himeno Takahashi²,
Mime Nagai²,
Hikari Sugawa² &
Ryoji Nagai ORCID: orcid.org/0000-0003-0763-0067^1,2

Amino Acids volume 54, pages 653–661 (2022)Cite this article

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Abstract

Cysteine is non-enzymatically modified by fumarate, which is an intermediate of the tricarboxylic acid cycle, leading to the formation of S-(2-succinyl)cysteine (2SC). Post-translational modification of physiological proteins by fumarate causes enzyme dysfunction. The aim of the study was to evaluate the changes in 2SC accumulation in physiological tissues associated with aging. Brain, liver, kidney, and serum samples were collected from 4-, 12-, and 96-week-old male C57BL/6J mice, and the level of 2SC was determined by liquid chromatography–tandem mass spectrometry (LC–MS/MS) after pretreatment, including delipidation, protein precipitation, and hydrolysis using hydrochloric acid. The 2SC level in the brain was higher than that in other tissues, and its accumulation significantly increased with age. Similarly, N^ε-(carboxymethyl)lysine levels, an advanced glycation end-products (AGEs) that accumulates in tissues in an age-dependent manner, was found to be increased in the brain and kidneys of elderly mice. Accumulation of N^δ-(5-hydro-5-methyl-4-imidazolone-2-yl)-ornithine increased significantly with age, but only in the kidneys. The fumarate content in the brain was similar to that in the liver and kidney at 4 and 12 weeks of age. Furthermore, fumarate contents increased in the liver and kidney at 96 weeks of age, whereas its level did not change in the brain. Our results demonstrated that the changes in 2SC and AGEs levels in tissues reflected differing metabolism and enhanced oxidative stress in each organ; in particular, the metabolism in the brain and kidneys is highly affected by aging.

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Abbreviations

AGEs:: Advanced glycation end-products
BCA:: Bicinchoninic acid
CEL:: N^ε-(carboxyethyl)lysine
CML:: N^ε-(carboxymethyl)lysine
GLO1:: Glyoxalase 1
HCl:: Hydrochloric acid
LC–MS/MS:: Liquid chromatography–tandem mass spectrometry
MG:: Methylglyoxal
MG-H1:: N^δ-(5-hydro-5-methyl-4-imidazolone-2-yl)-ornithine
NADH:: Reduced nicotinamide adenine dinucleotide
NAD⁺ :: Oxidized nicotinamide adenine dinucleotide
NaOH:: Sodium hydroxide
PTM:: Post-translational modification
SD:: Standard deviation
TCA:: Trichloroacetic acid
2SC:: S-(2-Succinyl)cysteine

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Acknowledgements

We are grateful to Jun-ichi Shirakawa and Hikari Satou from our laboratory for discussion about pretreatment of samples before analysis. The authors also thank Editage for editing the manuscript and providing suggestions.

Funding

This work was supported by a JSPS KAKENHI grant (grant no. 20K05895) provided to Ryoji Nagai. This work was also supported in part by the Japan Science and Technology Agency, Adaptable and Seamless Technology Transfer Program through Target-Driven R and D (number AS3015118U).

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Laboratory of Food and Regulation Biology, Graduate School of Bioscience, Tokai University, Kumamoto, Japan
Nana Katsuta & Ryoji Nagai
Laboratory of Food and Regulation Biology, Department of Bioscience, School of Agriculture, Tokai University, Toroku 9-1-1, Higashi-ku, Kumamoto City, Kumamoto, 862-8652, Japan
Himeno Takahashi, Mime Nagai, Hikari Sugawa & Ryoji Nagai

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Nana Katsuta
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Himeno Takahashi
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Mime Nagai
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Hikari Sugawa
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Ryoji Nagai
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Contributions

NK: Master of agriculture made principal contributions to study design and to the acquisition, analysis, and interpretation of results. She wrote the greater part of this article. HT: took part in animal experiments. MN: took part in the acquisition of data by measuring 2SC and AGEs. HS: took part in animal experiments, acquisition of data, and interpretation of results. RN: contributed to study design, measuring 2SC and AGEs, and interpretation of results.

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Correspondence to Ryoji Nagai.

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Animal experiments were performed with the approval of the ethics review committee of Tokai University for animal experimentation (approval number 211019).

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726_2022_3130_MOESM1_ESM.tif

Supplementary file1 (TIF 94 KB) A scheme of pretreatment procedures for the measurement of 2SC and AGEs. (A) A scheme of pretreatment for measurement of 2SC levels in tissues. (B) A scheme of pretreatment of tissues for measurement of AGEs levels.

726_2022_3130_MOESM2_ESM.tif

Supplementary file2 (TIF 141 KB) Changes in AGEs levels normalized by protein. CML levels in the (A) brain, (B) liver, and (C) kidneys was measured by LC-MS/MS and normalized to the protein content of each tissue. MG-H1 levels in the (D) brain, (E) liver, and (F) kidneys was measured by LC-MS/MS and normalized to the protein content of each tissue. Comparison of AGEs contents at 4, 12, and 96 weeks of age is presented. Graph that shows the data as jittered dots. The summary of the data is shown as horizontal line indicating the median (n=6, ∗p < 0.05).

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Katsuta, N., Takahashi, H., Nagai, M. et al. Changes in S-(2-succinyl)cysteine and advanced glycation end-products levels in mouse tissues associated with aging. Amino Acids 54, 653–661 (2022). https://doi.org/10.1007/s00726-022-03130-y

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Received: 01 September 2021
Accepted: 15 January 2022
Published: 15 February 2022
Issue Date: April 2022
DOI: https://doi.org/10.1007/s00726-022-03130-y

Keywords

S-(2-succinyl)cysteine
Advanced glycation end-products
Aging-related changes
Liquid chromatography–tandem mass spectrometry
Brain metabolite levels

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